Compositions, kits, and methods relating to the human FEZ1 gene, a novel tumor suppressor gene

Abstract

The invention relates to isolated polynucleotides homologous with a portion of one strand of the human tumor suppressor gene, FEZ1, and to the tumor suppressor protein encoded thereby, Fez1. The polynucleotides are useful, for example, as probes, primers, portions of expression vectors, and the like. The invention also includes diagnostic, therapeutic, cell proliferation enhancement, and screening methods which involve these polynucleotides and protein. The invention further includes kits useful for performing the methods of the invention.

1. A kit for amplifying a portion of a human FEZ1 gene, the kit comprising a first isolated polynucleotide and a second isolated polynucleotide, wherein the first isolated polynucleotide comprises a portion which anneals with high stringency with at least twenty consecutive nucleotide residues of the coding strand of SEQ ID NO: 1, and wherein the second isolated polynucleotide comprises a portion which anneals with high stringency with at least twenty consecutive nucleotide residues of the non-coding strand of SEQ ID NO:1. 
     
     
       2. A kit for amplifying a portion of a cDNA generated from a transcript of a human FEZ1 gene, the kit comprising a first isolated polynucleotide and a second isolated polynucleotide, wherein a portion of the first isolated polynucleotide anneals with high stringency with at least twenty consecutive nucleotide residues of the coding strand of SEQ ID NO: 1, and wherein a portion of the second isolated polynucleotide anneals with high stringency with at least twenty consecutive nucleotide residues of the non-coding strand of SEQ ID NO: 1. 
     
     
       3. An isolated polynucleotide comprising a sequence that anneals under conditions of high stringency to a nucleic acid having the sequence of:
 i) SEQ ID NO: 1; 
 ii) the complement of SEQ ID NO: 1; 
 iii) SEQ ID NO: 2; 
 iv) the complement of SEQ ID NO: 2; 
 v) SEQ ID NO: 3; and/or 
 vi) the complement of SEQ ID NO: 3; 
 wherein said conditions of high stringency comprise hybridizing said isolated polynucleotide in 0.015 M NaCl, 1.5 mM sodium citrate, and 0.1% (w/v) SDS at 50° C., with washes at 42° C. in 0.2×SSC and 0.1% (w/v) SDS; and wherein said isolated polynucleotide encodes a protein that binds to a compound selected from the group consisting of an amino-terminal 40 KDa fragment of Fez1, tubulin, EF1-γ, and an amino terminal 153-amino acid fragment of EF1-γ. 
 
     
     
       4. The isolated polynucleotide of  claim 3 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of SEQ ID NO: 1. 
     
     
       5. The isolated polynucleotide of  claim 3 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of the complement of SEQ ID NO: 1. 
     
     
       6. The isolated polynucleotide of  claim 3 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of SEQ ID NO: 2. 
     
     
       7. The isolated polynucleotide of  claim 3 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of the complement of SEQ ID NO: 2. 
     
     
       8. The isolated polynucleotide of  claim 3 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of SEQ ID NO: 3. 
     
     
       9. The isolated polynucleotide of  claim 3 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of the complement of SEQ ID NO: 3. 
     
     
       10. The isolated polynucleotide of  claim 3  further comprising a promoter operably linked to said sequence. 
     
     
       11. The isolated polynucleotide of  claim 10 , wherein said promoter is selected from the group consisting of a constitutive promoter, an inducible promoter and a tissue specific promoter. 
     
     
       12. The isolated polynucleotide of  claim 3 , wherein said protein inhibits tubulin polymerization. 
     
     
       13. The isolated polynucleotide of  claim 3 , wherein said protein inhibits cellular proliferation. 
     
     
       14. The isolated polynucleotide of  claim 3 , wherein said protein inhibits tumorigenesis. 
     
     
       15. A nucleic acid vector comprising the isolated polynucleotide of  claim 3 . 
     
     
       16. The nucleic acid vector of  claim 15  selected from the group consisting of a plasmid, an expression vector and a viral vector. 
     
     
       17. An isolated cell comprising the nucleic acid vector of  claim 15 . 
     
     
       18. An isolated polynucleotide comprising a sequence that anneals under conditions of high stringency to a nucleic acid having the sequence of
 i) SEQ ID NO: 1; 
 ii) the complement of SEQ ID NO: 1; 
 iii) SEQ ID NO: 2; 
 iv) the complement of SEQ ID NO: 2; 
 v) SEQ ID NO: 3; and/or 
 vi) the complement of SEQ ID NO: 3; 
 wherein said conditions of high stringency comprise hybridizing said isolated polynucleotide in wherein said conditions of high stringency comprise hybridizing said isolated polynucleotide in 0.015 M NaCl, 1.5 mM sodium citrate, and 0.1% (w/v) SDS at 50° C., with washes at 42° C. in 0.2×SSC and 0.1% (w/v) SDS; and wherein said isolated polynucleotide encodes a protein that has an activity selected from the group consisting of inhibiting cellular proliferation and tumor suppression. 
 
     
     
       19. The isolated polynucleotide of  claim 18 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of SEQ ID NO: 1. 
     
     
       20. The isolated polynucleotide of  claim 18 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of the complement of SEQ ID NO: 1. 
     
     
       21. The isolated polynucleotide of  claim 18 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of SEQ ID NO: 2. 
     
     
       22. The isolated polynucleotide of  claim 18 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of the complement of SEQ ID NO: 2. 
     
     
       23. The isolated polynucleotide of  claim 18 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of SEQ ID NO: 3. 
     
     
       24. The isolated polynucleotide of  claim 18 , wherein said isolated polynucleotide anneals under conditions of high stringency to a nucleic acid having the sequence of the complement of SEQ ID NO: 3. 
     
     
       25. The isolated polynucleotide of  claim 18  further comprising a promoter operably linked to said sequence. 
     
     
       26. The isolated polynucleotide of  claim 25 , wherein said promoter is selected from the group consisting of a constitutive promoter, an inducible promoter and a tissue specific promoter. 
     
     
       27. A nucleic acid vector comprising the isolated polynucleotide of  claim 18 . 
     
     
       28. The nucleic acid vector of  claim 27  selected from the group consisting of a plasmid, an expression vector and a viral vector. 
     
     
       29. An isolated cell comprising the nucleic acid vector of  claim 27 . 
     
     
       30. An isolated polynucleotide comprising a nucleotide sequence that encodes a protein comprising the amino acid sequence of SEQ ID NO: 4. 
     
     
       31. The isolated polynucleotide of  claim 30  wherein the isolated polynucleotide encodes the polypeptide of SEQ ID NO: 4. 
     
     
       32. The isolated polynucleotide of  claim 30  further comprising a promoter operably linked to said sequence. 
     
     
       33. The isolated polynucleotide of  claim 32 , wherein said promoter is selected from the group consisting of a constitutive promoter, an inducible promoter and a tissue specific promoter. 
     
     
       34. A nucleic acid vector comprising the isolated polynucleotide of  claim 30 . 
     
     
       35. The nucleic acid vector of  claim 34  selected from the group consisting of a plasmid, an expression vector and a viral vector. 
     
     
       36. An isolated cell comprising the nucleic acid vector of  claim 34 . 
     
     
       37. An isolated polynucleotide comprising a nucleotide sequence that encodes a protein comprising an amino terminal 40 KDa fragment of the sequence of SEQ ID NO: 4. 
     
     
       38. The isolated polynucleotide of  claim 37  wherein the isolated polynucleotide encodes the amino terminal 40 KDa fragment of the sequence of SEQ ID NO: 4. 
     
     
       39. The isolated polynucleotide of  claim 37  further comprising a promoter operably linked to said sequence. 
     
     
       40. The isolated polynucleotide of  claim 39 , wherein said promoter is selected from the group consisting of a constitutive promoter, an inducible promoter and a tissue specific promoter. 
     
     
       41. A nucleic acid vector comprising the isolated polynucleotide of  claim 37 . 
     
     
       42. The nucleic acid vector of  claim 41  selected from the group consisting of a plasmid, an expression vector and a viral vector. 
     
     
       43. An isolated cell comprising the nucleic acid vector of  claim 41 .